Zinc finger is one of DNA-binding motives, as well as the helix-turn-helix motif and the leucine zipper motif. It has two cysteine residues in the amino terminus region and two histidine residues in the carboxyl terminus region, and takes a three dimensional structure in which zinc (Zn) coordinates with these residues. Since zinc finger has an extremely strong DNA-binding ability, artificial DNA-binding proteins that utilize the motif to strongly bind to DNA (henceforth also referred to as “AZP” in this specification) have been proposed, and there have been reported AZPs designed so that they can recognize a specific nucleotide sequence by using the nondegenerate recognition code table (Japanese Patent Unexamined Publication (KOHYO) No. 2004-519211; Biochemistry, 41, pp. 7074-7081, 2002).
One zinc finger motif can recognize 3 or 4 bps and bind to the base pairs, and by connecting zinc fingers with a peptide linker, length of nucleotide sequence desired to be specifically bound by zinc fingers can be controlled. The fourth recognition nucleotide sequence of the zinc finger motif is an antisense strand, and overlaps with the first recognition nucleotide sequence of the following zinc finger motif, and therefore, N zinc finger motifs recognize a nucleotide sequence of 3N+1 bps, and bind thereto (see FIG. 1).
It has been reported that infection of plant DNA viruses can be controlled by using the AZP (J. Virology, 79, pp. 2614-2619, 2005). This publication reports infection-controlling effect of AZP for the plant DNA virus, beet severe curly top virus (BSCTV), in Arabidopsis thaliana. In this method, a means of inhibition by AZP is applied against the binding of a replication protein (Rep) to the Rep binding site (direct repeats) on the replication origin, which binding is required for the start of virus replication, and this method includes design of AZP to inhibit virus replication so that the AZP has a DNA-binding ability higher than that of Rep on the basis of the direct repeats of the replication origin. However, since the replication origin has a virus-specific nucleotide sequence, this method, including the block of the direct repeats of Rep by the AZP, has a problem that use of each different AZP is required against each of various plant viruses. From this point of view, it is desired to provide a means for achieving infection-controlling effect for various plant viruses with a single AZP.
Tomato yellow leaf curl is a viral disease of tomato plants and is induced by infection of the tomato yellow leaf curl virus (TYLCV). When the tomato yellow leaf curl is developed, green color of fresh leaves fades and yellows, leaves curl and then shrink, and the whole plant shows dwarfing, which results in arrest of growth. The tomato yellow leaf curl has caused serious damages in the Mediterranean sea coast, Africa, Middle and Near East, Asia, Central and South America, and the like. There are many isolated strains of TYLCV, and the TYLCV Israel strain (fulminant type, Nagasaki strain and Tosa strain), the TYLCV mild strain (mild type, Shizuoka strain and Aichi strain), and the like have so far been reported in Japan.
TYLCV belongs to the family Geminiviridae, and geminivirus is a generic name of viruses having one or two single-stranded cyclic DNAs that infect plants. Geminiviruses include various plant viruses, such as potato yellow mosaic virus and bean golden mosaic virus. If a means for inhibition can be provided against virus replication targeting a nucleotide sequence highly conserved in the geminiviruses, it is expected that TYLCV infection as well as infection with various plant viruses can be effectively controlled. Although the method disclosed in International Patent Publication WO2004/101798 and the like are known as a method for preparing a transformed plant having sustainable resistance against geminivirus, the approach thereof is completely different from that of the present invention.